Tariffs on cars, large and small trucks, car parts and steel will be increased to 50 per cent, Trump said on his Truth Social platform.
"Build in the US and there are ZERO TARIFFS," Trump wrote.
According to Canadian government figures, the US currently imposes tariffs of 25 per cent on imported cars, trucks and related parts under Section 232 of the Trade Expansion Act of 1962.
Tariffs of between 10 per cent and 50 per cent are also imposed on imports of steel, aluminium and copper.
Trump again justified his threat by accusing Canada of taking advantage of the US.
"Canada has been ripping off the United States of America for years," he wrote, while also claiming that the US did not depend on trade with its neighbour.
"WE DON'T NEED CANADA, THEY NEED US!" Trump wrote.
Negotiators from the two countries had held talks into Friday night in an effort to reach an agreement and avert tariffs threatened by Trump.
After the negotiations collapsed, tariffs of 50 per cent on a range of Canadian goods came into force overnight.
The measures affect Canadian goods worth about $US20 billion ($A28 billion), equivalent to about five per cent of total imports, including hockey sticks, furniture, honey and wine.
Canada vowed reciprocal "dollar-for-dollar" tariffs, with Prime Minister Mark Carney saying these would apply from September 8.
